The focus of this work has been to understand the molecular details that control initial steps in the recognition of cells infected with pathogens such as viruses by cells of the innate and adaptive immune systems. Understanding the function, mechanism, structure, and evolution of the interaction of virus-encoded molecules recognized by the immune system can lead not only to a deeper understanding of molecular interactions in general and of cell-cell interactions in the immune system, but also may lead to rational approaches to intervention in virus infection and neoplasia. In particular, we study representative members of the large family of major histocompatibility complex (MHC)-encoded molecules from a biophysical and structural perspective. We are interested in how MHC-I molecules interact with receptors on natural killer (NK) cells and on T lymphocytes through their NK and T cell receptors, respectively. Large DNA viruses of the herpesvirus family produce proteins that mimic host MHC-I molecules as part of their immunoevasive strategy, and we have directed our efforts to understand the function, cellular expression, and structure of a set of these MHC-I (referred to as MHC-Iv) molecules encoded by the mouse cytomegalovirus (mCMV). We have analyzed the expression and binding of several members of the MHC-Iv family, in particular the molecules, m144, m152, and m153. In earlier studies, we determined the structures of the MHC-Iv molecules, m144, m152, and m153. In a recent collaboration with Drs. Oscar Aguilar and James Carlyle (manuscript submitted) at the University of Toronto, we have demonstrated that recombinant m153, multimerized as a staining agent, specifically recognizes molecules expressed on the surface of a dendritic cell line, DC2.4, as well as on an ILC3-like NK cell, MNK-3. This finding complements our collaborators finding that m153 regulates the cell surface level of an NK cell ligand, Clr-b. In addition to our studies of the viral MHC-like molecules, we have recently addressed another set of the putative mCMV immunoevasins, the m04 family. This family, that includes m02, m04, and m06 seems to have distinct mechanisms of action. m04 accompanies the MHC-I molecule to the cell surface, and m06 directs MHC-I molecules to an endosomal/lysosomal pathway. We have successfully engineered m04 and examined its binding to MHC-I to quantify this interaction. We previously determined the structure of m04 in solution using NMR in collaboration with Drs. Nik Sgourakis and Ad Bax. NMR structure determination is based on determination of multidimensional spectra that allow assignment of molecular restraints to various interatomic distances within the molecule. Proceeding from these earlier studies, we are now complementing them with cellular imaging experiments designed to examine the precise intermediates of MHC-I folding that proceed within the cell. We have established that m06 routes MHC-I molecules to the endosomal/lysosomal pathway, thus contributing to the downregulation of cell surface MHC-I. Preliminary screening of molecular complexes of the m06 protein bound to the MHC molecule H2-Ld indicate that diffraction quality crystals should be achievable to allow definitive structural determination of the unique mechanism of m06 evasive function. Two large scale reviews of the comparative molecular structures of MHC molecules, host MHC-like molecules, and viral MHC-like molecules (Natarajan et al, 2018; and Jiang et al, 2019, in press) indicate that the general ability of molecules of the MHC family to exhibit both specificity and degeneracy lies in regions of molecular flexibility, related to either peptide or ligand interaction sites. Recent experiments from other labs have identified a novel immunoevasin encoded by the Molluscum contagiosum virus, MC80, that impedes antigen presentation by interacting with tapasin in the protein loading complex, and directs tapasin to an endoplasmic reticulum degradative pathway. We are currently expressing the MC80 protein for structural and binding studies, and expect that this basic knowledge will both enhance our understanding of the MHC peptide loading pathway as well as of the many avenues of viral immunoevasion.